Guanine nucleotide-binding regulatory proteins (GNPs) regulate cell growth and function through the transduction of receptor-initiated signals across cell membranes. Go is a member of the GNP family whose physiologic function has not been defined. Go exists in the brain in high quantities (as much as 1% of membrane protein), and interacts with rhodopsin and muscarinic receptors. Like other known GNPs, Go is a heterotrimer of alpha, beta, and gamma subunits. A cDNA clone (lambda GO9) containing the complete coding sequence for the alpha subunit of the bovine retina Go (Go alpha) (Van Meurs et al., Proc Natl Acad Sci USA 84:3107, 1987) was used to probe bovine messenger RNA. Three different sizes of Go alpha mRNA were found in bovine retina and two in brain. To determine the molecular basis of the heterogeneity in Go alpha mRNA transcripts, lambda GO9 was used as a probe to obtain related cDNAs from a bovine retinal library. Four clones were isolated and sequenced. Their 3' untranslated regions (UTRs) differ markedly from that of the original lambda GO9, containing in the divergent area a unique Hind III restriction site not present in lambda GO9. Northern analysis was carried out using 48-base oligonucleotides complementary to unique sequences in the two types of 3'UTRs. The probe complementary to the Hind III site-positive clones detected 2.0 and 4.0 kb Go alpha mRNA species, whereas the probe specific for the 3' UTR of lambda GO9 detected only 3.0 and 4.0 kb species. The 2.0 kb mRNA was present in retina, but not in brain. These specific probes to the alternate 3' UTRs should provide tools for studying tissue-specific Go alpha expression.